Vangl2 Gene Research Articles

A method for TAT-Cre recombinase-mediated floxed allele modification in ex vivo tissue slices.

Precision-cut lung slices (PCLS) are used for a variety of applications. However, methods to manipulate genes in PCLS are currently limited. We developed a new method, TAT-Cre recombinase-mediated floxed allele modification in tissue slices (TReATS), to induce highly effective and temporally controlled gene deletion or activation in ex vivo PCLS. Treatment of PCLS from Rosa26-flox-stop-flox-EYFP mice with cell-permeant TAT-Cre recombinase induced ubiquitous EYFP protein expression, indicating successful Cre-mediated excision of the upstream loxP-flanked stop sequence. Quantitative real-time PCR confirmed induction of EYFP. We successfully replicated the TReATS method in PCLS from Vangl2flox/flox mice, leading to the deletion of loxP-flanked exon 4 of the Vangl2 gene. Cre-treated Vangl2flox/flox PCLS exhibited cytoskeletal abnormalities, a known phenotype caused by VANGL2 dysfunction. We report a new method that bypasses conventional Cre-Lox breeding, allowing rapid and highly effective gene manipulation in ex vivo tissue models.

The loop-tail mouse model displays open and closed caudal neural tube defects.

Neural tube defects (NTDs) are the second most common cause of congenital malformations and are often studied in animal models. Loop-tail (Lp) mice carry a mutation in the Vangl2 gene, a member of the Wnt-planar cell polarity pathway. In Vangl2+/Lp embryos, the mutation induces a failure in the completion of caudal neural tube closure, but only a small percentage of embryos develop open spina bifida. Here, we show that the majority of Vangl2+/Lp embryos developed caudal closed NTDs and presented cellular aggregates that may facilitate the sealing of these defects. The cellular aggregates expressed neural crest cell markers and, using these as a readout, we describe a systematic method to assess the severity of the neural tube dorsal fusion failure. We observed that this defect worsened in combination with other NTD mutants, Daam1 and Grhl3. Besides, we found that in Vangl2+/Lp embryos, these NTDs were resistant to maternal folic acid and inositol supplementation. Loop-tail mice provide a useful model for research on the molecular interactions involved in the development of open and closed NTDs and for the design of prevention strategies for these diseases.

Dihydroartemisinin Attenuates Hypoxic Pulmonary Hypertension via the Downregulation of miR-335 Targeting Vangl2.

Dihydroartemisinin (DHA) is a traditional antimalarial drug. DHA plays a crucial role in preventing pulmonary hypertension (PH); however, its regulatory function on microRNAs (miRNAs) in PH remains unclear. This study aimed to investigate whether DHA exerts its protective functions by regulating miR-335 in PH. Hypoxia-induced PH models were induced both in vitro and in vivo. Mice were treated with various concentrations of DHA, and pulmonary arterial smooth muscle cells (PASMCs) were treated with DHA, miR-335 inhibitor, miR-335 mimic, or Van Gogh-like 2 (Vangl2) plasmid. The expression of miR-335 and Vangl2, pulmonary arterial remodeling index; right ventricular hypertrophy index; and proliferation and migration indexes were measured. DHA improved pulmonary vascular remodeling and alleviated PH in vivo. miRNA sequencing and real-time PCR results further show that the increase in hypoxia-induced miR-335 was avoided by DHA administration, and miR-335 increased the hypoxia-induced PASMC proliferation and migration. MiRNA databases and dual-luciferase reporter assay show that miR-335 directly targets Vangl2, and Vangl2 decreased the hypoxia-induced PASMC proliferation and migration. The miR-335 inhibitor failed to inhibit hypoxia-induced proliferation and migration upregulation in Vangl2 knockdown PASMCs, and the effect of DHA can be blocked by miR-335 upregulation. In hypoxic PH, MiR-335 is increased, whereas Vangl2 is decreased. MiR-335 can significantly promote the hypoxia-induced proliferation and migration of PASMCs by targeting the Vangl2 gene. DHA effectively reverses the hypoxia-induced upregulation of miR-335 expression, avoiding the miR-335-mediated downregulation of Vangl2 and thereby promoting the expression of Vangl2 to prevent PH.

Vangl2, a Core Component of the WNT/PCP Pathway, Regulates Adult Hippocampal Neurogenesis and Age-Related Decline in Cognitive Flexibility.

Decline in episodic memory is one of the hallmarks of aging and represents one of the most important health problems facing Western societies. A key structure in episodic memory is the hippocampal formation and the dentate gyrus in particular, as the continuous production of new dentate granule neurons in this brain region was found to play a crucial role in memory and age-related decline in memory. As such, understanding the molecular processes that regulate the relationship between adult neurogenesis and aging of memory function holds great therapeutic potential. Recently, we found that Vang-Gogh like 2 (Vangl2), a core component of the Planar Cell Polarity (PCP) signaling pathway, is enriched in the dentate gyrus of adult mice. In this context, we sought to evaluate the involvement of this member of the Wnt/PCP pathway in both adult neurogenesis and memory abilities in adult and middle-aged mice. Using a heterozygous mouse model carrying a dominant-negative mutation in the Vangl2 gene, called Looptail (Vangl2Lp), we show that alteration in Vangl2 expression decreases the survival of adult-born granule cells and advances the onset of a decrease in cognitive flexibility. The inability of mutant mice to erase old irrelevant information to the benefit of new relevant ones highlights a key role of Vangl2 in interference-based forgetting. Taken together, our findings show that Vangl2 activity may constitute an interesting target to prevent age-related decline in hippocampal plasticity and memory.

Cell non-autonomy amplifies disruption of neurulation by mosaic Vangl2 deletion in mice

Post-zygotic mutations that generate tissue mosaicism are increasingly associated with severe congenital defects, including those arising from failed neural tube closure. Here we report that neural fold elevation during mouse spinal neurulation is vulnerable to deletion of the VANGL planar cell polarity protein 2 (Vangl2) gene in as few as 16% of neuroepithelial cells. Vangl2-deleted cells are typically dispersed throughout the neuroepithelium, and each non-autonomously prevents apical constriction by an average of five Vangl2-replete neighbours. This inhibition of apical constriction involves diminished myosin-II localisation on neighbour cell borders and shortening of basally-extending microtubule tails, which are known to facilitate apical constriction. Vangl2-deleted neuroepithelial cells themselves continue to apically constrict and preferentially recruit myosin-II to their apical cell cortex rather than to apical cap localisations. Such non-autonomous effects can explain how post-zygotic mutations affecting a minority of cells can cause catastrophic failure of morphogenesis leading to clinically important birth defects.

Differential role of planar cell polarity gene Vangl2 in embryonic and adult mammalian kidneys.

Planar cell polarity (PCP) pathway is crucial for tissue morphogenesis. Mutations in PCP genes cause multi-organ anomalies including dysplastic kidneys. Defective PCP signaling was postulated to contribute to cystogenesis in polycystic kidney disease. This work was undertaken to elucidate the role of the key PCP gene, Vangl2, in embryonic and postnatal renal tubules and ascertain whether its loss contributes to cyst formation and defective tubular function in mature animals. We generated mice with ubiquitous and collecting duct-restricted excision of Vangl2. We analyzed renal tubules in mutant and control mice at embryonic day E17.5 and postnatal days P1, P7, P30, P90, 6- and 9-month old animals. The collecting duct functions were analyzed in young and adult mutant and control mice. Loss of Vangl2 leads to profound tubular dilatation and microcysts in embryonic kidneys. Mechanistically, these abnormalities are caused by defective convergent extension (larger tubular cross-sectional area) and apical constriction (cuboidal cell shape and a reduction of activated actomyosin at the luminal surface). However, the embryonic tubule defects were rapidly resolved by Vangl2-independent mechanisms after birth. Normal collecting duct architecture and functions were found in young and mature animals. During embryogenesis, Vangl2 controls tubular size via convergent extension and apical constriction. However, rapidly after birth, PCP-dependent control of tubular size is switched to a PCP-independent regulatory mechanism. We conclude that loss of the Vangl2 gene is dispensable for tubular elongation and maintenance postnatally. It does not lead to cyst formation and is unlikely to contribute to polycystic kidney disease.

Role of Core Planar Cell Polarity Vangl2 Gene in the Renal Tubule Development in Mice

Background: Polycystic kidney disease (PKD) is a common kidney disease that affects the development and maintenance of renal tubules, leads to cyst formation, and often progresses to end-stage kidney disease. It has been postulated that defective planar cell polarity (PCP) signaling contributes to initiation of cyst formation in PKD via controlling both convergent extension (CE, a process of directional cell movements) and oriented cell division (OCD, a process of directional cell divisions during tubular elongation post-natally). Indeed, mutations of the key PCP gene, Van Gogh-like 2 (Vangl2), lead to abnormal renal tubules in murine embryonic kidneys, correlating with the original postulate. Methods: In order to further understand the influence of the Vangl2 gene on renal morphogenesis and cystogenesis, control and Vangl2 mutant embryos—as well as post-natal Vangl2 mice with conditional excision of the Vangl2 gene in renal collecting tubules—were generated, then analyzed using immunostaining and fluorescence microscopy. Results: Our results show that Vangl2 plays a role in CE and apical constriction (AC) during embryonic stage of tubulogenesis. Compared to control animals, mutant Vangl2Δ/Δ and conditional Vangl2Δ/CD embryos displayed: i) a significant dilation in the diameter of renal tubules seen as an increased tubule cross-section area and a larger number of cells per cross-section; and ii) changes in cell shape indicative of defective AC. Surprisingly, post-natal mice showed virtually no difference in any of these aspects comparing to control mice, suggesting that other pathways may compensate for the lack of PCP signaling in maintenance of the tubule architecture. Limitations: a) The analysis of the renal tubules at the specific time points does not account for the dynamics of tubular movement and growth in real time; b) a mechanistic and morphological distinction between mice and humans may exist in the renal collecting duct tubules, pertaining to the Vangl2 gene’s influence in the PCP pathway; and c) the degree of mosaicism resulting from the gene excision by Cre-recombinase may correlate with the severity of the phenotype. Conclusion: We conclude that the PCP pathway is required for normal tubule development during embryogenesis. Our results, however, indicate that the cystogenesis seen in PKD postnatally may not be directly attributed to the disrupted PCP signaling, and requires the derangement of additional pathways.

The non-canonical Wnt-PCP pathway shapes the mouse caudal neural plate.

The last stage of neural tube (NT) formation involves closure of the caudal neural plate (NP), an embryonic structure formed by neuromesodermal progenitors and newly differentiated cells that becomes incorporated into the NT. Here, we show in mouse that, as cell specification progresses, neuromesodermal progenitors and their progeny undergo significant changes in shape prior to their incorporation into the NT. The caudo-rostral progression towards differentiation is coupled to a gradual reliance on a unique combination of complex mechanisms that drive tissue folding, involving pulses of apical actomyosin contraction and planar polarised cell rearrangements, all of which are regulated by the Wnt-PCP pathway. Indeed, when this pathway is disrupted, either chemically or genetically, the polarisation and morphology of cells within the entire caudal NP is disturbed, producing delays in NT closure. The most severe disruptions of this pathway prevent caudal NT closure and result in spina bifida. In addition, a decrease in Vangl2 gene dosage also appears to promote more rapid progression towards a neural fate, but not the specification of more neural cells.

Deficiency of the planar cell polarity protein Vangl2 in podocytes affects glomerular morphogenesis and increases susceptibility to injury.

The planar cell polarity (PCP) signaling pathway is crucial for tissue morphogenesis. Van Gogh-like protein 2 (Vangl2) is central in the PCP pathway; in mice, Vangl2 loss is embryonically lethal because of neural tube defects, and mutations in Vangl2 are associated with human neural tube defects. In the kidney, PCP signaling may be important for tubular morphogenesis and organization of glomerular epithelial cells (podocytes) along the glomerular basement membrane. Podocyte cell protrusions (foot processes) are critical for glomerular permselectivity; loss of foot process architecture results in proteinuria and FSGS. Previously, we showed a profound effect of PCP signaling on podocyte shape, actin rearrangement, cell motility, and nephrin endocytosis. To test our hypothesis that the PCP pathway is involved in glomerular development and function and circumvent lethality of the ubiquitous Vangl2 mutation in the Looptail mouse, we generated a mouse model with a podocyte-specific ablation of the Vangl2 gene. We report here that podocyte-specific deletion of Vangl2 leads to glomerular maturation defects in fetal kidneys. In adult mice, we detected significantly smaller glomeruli, but it did not affect glomerular permselectivity in aging animals. However, in the context of glomerular injury induced by injection of antiglomerular basement membrane antibody, deletion of Vangl2 resulted in exacerbation of injury and accelerated progression to chronic segmental and global glomerular sclerosis. Our results indicate that Vangl2 function in podocytes is important for glomerular development and protects against glomerular injury in adult animals.

Postnatal Refinement of Auditory Hair Cell Planar Polarity Deficits Occurs in the Absence of Vangl2

The distinctive planar polarity of auditory hair cells is evident in the polarized organization of the stereociliary bundle. Mutations in the core planar cell polarity gene Van Gogh-like 2 (Vangl2) result in hair cells that fail to properly orient their stereociliary bundles along the mediolateral axis of the cochlea. The severity of this phenotype is graded along the length of the cochlea, similar to the hair cell differentiation gradient, suggesting that an active refinement process corrects planar polarity phenotypes in Vangl2 knock-out (KO) mice. Because Vangl2 gene deletions are lethal, Vangl2 conditional knock-outs (CKOs) were generated to test this hypothesis. When crossed with Pax2-Cre, Vangl2 is deleted from the inner ear, yielding planar polarity phenotypes similar to Vangl2 KOs at late embryonic stages except that Vangl2 CKO mice are viable and do not have craniorachischisis like Vangl2 KOs. Quantification of planar polarity deficits through postnatal development demonstrates the activity of a Vangl2-independent refinement process that rescues the planar polarity phenotype within 10 d of birth. In contrast, the Pax2-Cre;Vangl2 CKO has profound changes in the shape and distribution of outer pillar cell and Deiters' cell phalangeal processes that are not corrected during the period of planar polarity refinement. Auditory brainstem response analyses of adult mice show a 10-15 dB shift in auditory threshold, and distortion product otoacoustic emission measurements indicate that this mild hearing deficit is of cochlear origin. Together, these data demonstrate a Vangl2-independent refinement mechanism that actively reorients auditory stereociliary bundles and reveals an unexpected role of Vangl2 during supporting cell morphogenesis.

Loop-tail phenotype in heterozygous mice and neural tube defects in homozygous mice result from a nonsense mutation in the Vangl2 gene

N-ethyl-N-nitrosourea (ENU) is a powerful point mutagen that can generate random mutations. It has been used to generate mouse mutations to produce phenotypic models of human disease. Neural tube defects (NTD) are common birth defects in which the brain and/or spinal cord can be exposed; however, the mechanisms of these defects are poorly understood. Craniorachischisis is one type of NTD that bears a close resemblance to the phenotype of the loop-tail (Lp) mouse. Here we describe a C57BL/6J Lp mouse generated by ENU-induced mutagenesis. The mutation was mapped to the Vangl2 gene on chromosome 1, near markers D1Mit113 and D1Mit149. Sequence analysis of Vangl2 heterozygotes (Vangl2(m1Yzcm)/+) revealed a C/T transition mutation that resulted in substitution of a glutamine codon for a stop (nonsense) codon at position 449. The Vangl2 protein is involved in epithelium planar cell polarity. The predicted truncated protein would lack the PDZ-domain binding motif involved in protein-protein interaction; therefore, Vangl2(m1Yzcm) may be a loss-of-function mutant. Morphological and histological examination of homozygous mouse embryos revealed a neural tube closure defect that leads to craniorachischisis. This Vangl2(m1Yzcm) mouse represents a valuable model for the study of NTDs in humans.

Vangl2 and planar cell polarity signaling in cardiac development

Planar cell polarity pathway regulates cell polarity and polarized cell movements.Recent studies in mouse models have found mutations in several genes in this pathway and specifically in the Vangl2 gene,which results in abnormalities in cardiac development,suggesting Vangl2 and this pathway play an important role in heart development.This review mainly elucidates the mechanisms regulated by the Vangl2 gene and PCP pathway during outflow tract development and the formation of the coronary vasculature. Key words: Vangl2; Noncanonical Wnt/PCP pathway; Cardiac development; Congenital heart disease

Molecular Characterisation of Endogenous Vangl2/Vangl1 Heteromeric Protein Complexes

BackgroundMutations in the Planar Cell Polarity (PCP) core gene Vangl2 cause the most severe neural tube defects (NTD) in mice and humans. Genetic studies show that the Vangl2 gene genetically interacts with a close homologue Vangl1. How precisely Vangl2 and Vangl1 proteins interact and crosstalk has remained a difficult issue to address, with the main obstacle being the accurate discrimination of the two proteins, which share close sequence homology. Experimental evidence previously presented has been sparse and addressed with ectopically expressed proteins or with antibodies unable to biochemically discriminate Vangl1 from Vangl2, therefore giving rise to unclear results.Methodology and Main FindingsA highly specific monoclonal anti-Vangl2 antibody was generated and rigorously tested on both recombinant and extracted Vangl2 using surface plasmon resonance (SPR) analysis, western blot, and immunoprecipitation experiments. This antibody efficiently affinity-purified Vangl2 from cell lysates and allowed the unambiguous identification of endogenous Vangl2 by proteomic analysis. Vangl1 was also present in Vangl2 immunoprecipitates, establishing the first biochemical evidence for the existence of Vangl2/Vangl1 heterodimers at an endogenous level. Epitope-tagged Vangl2 and Vangl1 confirmed that both proteins interact and colocalize at the plasma membrane. The Vangl2 antibody is able to acutely assess differential expression levels of Vangl2 protein in culture cell lines, as corroborated with gene expression analysis. We characterised Vangl2 expression in the cochlea of homozygous and heterozygous Lp mutant mice bearing a point mutation within the C-terminal Vangl2 region that leads to profound PCP defects. Our antibody could detect much lower levels of Vangl2Lp protein in mutant mice compared to the wild type mice.ConclusionOur results provide an in-depth biochemical characterisation of the interaction observed between Vangl paralogues.

Disheveled mediated planar cell polarity signaling is required in the second heart field lineage for outflow tract morphogenesis

Disheveled (Dvl) is a key regulator of both the canonical Wnt and the planar cell polarity (PCP) pathway. Previous genetic studies in mice indicated that outflow tract (OFT) formation requires Dvl1 and 2, but it was unclear which pathway was involved and whether Dvl1/2-mediated signaling was required in the second heart field (SHF) or the cardiac neural crest (CNC) lineage, both of which are critical for OFT development. In this study, we used Dvl1/2 null mice and a set of Dvl2 BAC transgenes that function in a pathway-specific fashion to demonstrate that Dvl1/2-mediated PCP signaling is essential for OFT formation. Lineage-specific gene-ablation further indicated that Dvl1/2 function is dispensable in the CNC, but required in the SHF for OFT lengthening to promote cardiac looping. Mutating the core PCP gene Vangl2 and non-canonical Wnt gene Wnt5a recapitulated the OFT morphogenesis defects observed in Dvl1/2 mutants. Consistent with genetic interaction studies suggesting that Wnt5a signals through the PCP pathway, Dvl1/2 and Wnt5a mutants display aberrant cell packing and defective actin polymerization and filopodia formation specifically in SHF cells in the caudal splanchnic mesoderm (SpM), where Wnt5a and Dvl2 are co-expressed specifically. Our results reveal a critical role of PCP signaling in the SHF during early OFT lengthening and cardiac looping and suggest that a Wnt5a→ Dvl PCP signaling cascade may regulate actin polymerization and protrusive cell behavior in the caudal SpM to promote SHF deployment, OFT lengthening and cardiac looping.

Comparison of Phenotypes between Different vangl2 Mutants Demonstrates Dominant Effects of the Looptail Mutation during Hair Cell Development

Experiments utilizing the Looptail mutant mouse, which harbors a missense mutation in the vangl2 gene, have been essential for studies of planar polarity and linking the function of the core planar cell polarity proteins to other developmental signals. Originally described as having dominant phenotypic traits, the molecular interactions underlying the Looptail mutant phenotype are unclear because Vangl2 protein levels are significantly reduced or absent from mutant tissues. Here we introduce a vangl2 knockout mouse and directly compare the severity of the knockout and Looptail mutant phenotypes by intercrossing the two lines and assaying the planar polarity of inner ear hair cells. Overall the vangl2 knockout phenotype is milder than the phenotype of compound mutants carrying both the Looptail and vangl2 knockout alleles. In compound mutants a greater number of hair cells are affected and changes in the orientation of individual hair cells are greater when quantified. We further demonstrate in a heterologous cell system that the protein encoded by the Looptail mutation (Vangl2S464N) disrupts delivery of Vangl1 and Vangl2 proteins to the cell surface as a result of oligomer formation between Vangl1 and Vangl2S464N, or Vangl2 and Vangl2S464N, coupled to the intracellular retention of Vangl2S464N. As a result, Vangl1 protein is missing from the apical cell surface of vestibular hair cells in Looptail mutants, but is retained at the apical cell surface of hair cells in vangl2 knockouts. Similarly the distribution of Prickle-like2, a putative Vangl2 interacting protein, is differentially affected in the two mutant lines. In summary, we provide evidence for a direct physical interaction between Vangl1 and Vangl2 through a combination of in vitro and in vivo approaches and propose that this interaction underlies the dominant phenotypic traits associated with the Looptail mutation.

A novel hypomorphic Looptail allele at the planar cell polarity Vangl2 gene

Vangl2 forms part of the planar cell polarity signalling pathway and is the gene defective in the Looptail (Lp) mouse mutant. Two previously described alleles, Lp and Lp(m1Jus) , segregate in a semi-dominant fashion, with heterozygotes displaying the looped-tail appearance, while homozygotes show the neural tube defect called craniorachischisis. Here, we report a novel experimentally induced allele, Lp(m2Jus) , that carries a missense mutation, R259L, in Vangl2. This mutation was specific to the Lp phenotype and absent from both parental strains and 28 other inbred strains. Notably, this mutation segregates in a recessive manner with all heterozygotes appearing normal and 47% of homozygotes showing a looped-tail. Homozygous Lp(m2Jus) embryos showed spina bifida in 12%. Lp(m2Jus) genetically interacts with Lp with 77% of compound heterozygotes displaying craniorachischisis. Vangl2(R259L) behaved like the wild-type allele in overexpression and morpholino knockdown/rescue assays in zebrafish embryos. These data suggest that Lp(m2Jus) represents a new hypomorphic allele of Lp.

Molecular and Cellular Mechanisms Underlying Neural Tube Defects in the Loop-tail Mutant Mouse

Loop-tail (Lp) mice show a very severe neural tube defect (craniorachischisis) caused by mutations in the Vangl2 gene (D255E, S464N). Mammalian Vangl1 and Vangl2 are membrane proteins that play critical roles in development such as establishment of planar cell polarity (PCP) in epithelial layers and convergent extension movements during neurogenesis and cardiogenesis. Vangl proteins are thought to assemble with other PCP proteins (Dvl, Pk) to form membrane-bound PCP signaling complexes that provide polarity information to the cell. In the present study, we show that Vangl1 is expressed exclusively at the plasma membrane of transfected MDCK cells, where it is targeted to the basolateral membrane. Experiments with an inserted exofacial HA epitope indicate that the segment delimited by the predicted transmembrane domains 1 and 2 is exposed to the extracellular milieu. Comparative studies of the Lp-associated pathogenic mutation D255E indicate that the targeting of the mutant variant at the plasma membrane is greatly reduced; the mutant variant is predominantly retained intracellularly in endoplasmic reticulum (ER) vesicles colocalizing with the ER marker calreticulin. In addition, the D255E variant shows drastically reduced stability with a half-life of approximately 2 h, compared to >9 h for its wild type counterpart and is rapidly degraded in a proteasome-dependent and MG132 sensitive pathway. These findings highlight a critical role for D255 for normal folding and processing of Vangl proteins, with highly conservative substitutions not tolerated at that site. Our study provide an experimental framework for the analysis of human VANGL mutations recently identified in familial and sporadic cases of spina bifida.

Investigation of Van Gogh-like 2 mRNA regulation and localisation in response to nociception in the brain of adult common carp ( Cyprinus carpio)

The Van Gogh-like 2 ( vangl2) gene is typically associated with planar cell polarity pathways, which is essential for correct orientation of epithelial cells during development. The encoded protein of this gene is a transmembrane protein and is highly conserved through evolution. Van Gogh-like 2 was selected for further study on the basis of consistent regulation after a nociceptive stimulus in adult common carp and rainbow trout in a microarray study. An in situ hybridisation was conducted in the brain of mature common carp ( Cyprinus carpio), 1.5 and 3 h after a nociceptive stimulus comprising of an acetic acid injection to the lips of the fish and compared with a saline-injected control. The vangl2 gene was expressed in all brain regions, and particularly intensely in neurons of the telencephalon and in ependymal cells. In the cerebellum, a greater number ( P = 0.018) of Purkinje cells expressed vangl2 after nociception ( n = 7) compared with controls ( n = 5). This regulation opens the possibility that vangl2 is involved in nociceptive processing in the adult fish brain and may be a novel target for central nociception in vertebrates.

Mutation Analysis of the Vangl2 Coding Region Revealed No Common Cause for Tetralogy of Fallot

Vangl2 (Van Gogh-like 2) protein acts via non-canonical Wnt signalling to regulate polarized cell movements during development of the proximal outflow tract in vertebrate embryos. Recently, it has been shown that mutations of the Vangl2 gene cause aortic arch defects that are characteristic of the loop-tail (Lp) mouse and they have also became a strong candidate for causing congenital outflow tract defects in humans. Thus, in this study Tetralogy of Fallot (ToF), which comprises a group of syndromes that constitutes the most frequent cause of congenital cardiac outflow abnormalities in humans, was analysed for mutations within all coding regions of the Vangl2 gene. Based on direct sequencing data from a combination of 20 patients with ToF and 22 healthy people, three polymorphisms have been identified in exon 6 and exon 7 which do not change the amino acid sequence. It was concluded, therefore, that there is no specific mutation responsible for the ToF phenotype in the Vangl2 gene.

Tissue, cellular and sub-cellular localization of the Vangl2 protein during embryonic development: Effect of the Lp mutation

Loop-tail ( Lp) mice show a very severe neural tube defect, craniorachischisis, which is caused by mis-sense mutations in the Vangl2 gene. The membrane protein Vangl2 belongs to a highly conserved group of proteins that regulate planar polarity in certain epithelia, and that are also important for convergent extension movements during gastrulation and neurulation. A specific anti-Vangl2 antiserum was produced and used to examine the tissue, cell type, and sub-cellular localization of Vangl2 during embryogenesis. Vangl2 protein is expressed at high levels in the neural tube and shows a dynamic expression profile during neurulation. After neural tube closure, robust Vangl2 staining is detected in several neural and neurosensory tissues, including cerebral cortex, dorsal root ganglia, olfactory epithelium, retina, mechanosensory hair cells of the cochlea, and optic nerve. Vangl2 is also expressed during organogenesis in a number of tubular epithelia, including the bronchial tree, intestinal crypt/villus axis, and renal tubular segments derived from ureteric bud and from metanephric mesenchyme. Examination of Vangl2 localization in the neural tubes and cochleas of the normal and Lp/ Lp embryos shows disruption of normal membrane localization of Vangl2 in independent alleles at Lp ( Lp, Lp m1Jus ) as well as overall decrease in the expression level.